Nitrogenase (N2ase) is a metalloenzyme that mediates biological nitrogen fixation and is essential to life. As such, the study of nitrogenase attracts intense scrutiny among the biology and chemistry communities. Nonetheless, the mechanism by which nitrogenase enzymes promote the biological reduction of nitrogen under ambient conditions remains a fascinating and unsolved problem. The broad goal of our research is to evaluate the mechanisms by which a single iron site is able to mediate catalytic N2 reduction in synthetic model systems and, by extension, in biology. The proposed program is to design and study biomimetic Fe-NxHy model complexes to address this goal. Our experimental approach stresses functionally, rather than structurally, faithful models of the iron-molybdenum cofactor (FeMoco). Low molecular weight Fe-NxHy complexes will be developed to explore iron sites in low coordinate geometries that may accommodate dinitrogen and other NxHy functionalities. We posit these geometries as relevant to Fe-NxHy intermediates of the FeMoco. By analogy to the modes of Fe-mediated biocatalytic O2 reduction, two limiting single-site mechanisms are emphasized. The first is an alternating mechanism, where successive H-atom transfers (via H+/e- steps) occur at the distal and proximal N-atoms of the Fe-N?N subunit in an alternating fashion (e.g., Fe- N=NH ? Fe-NH=NH ? Fe-NH-NH2? Fe-NH2-NH2 ? Fe-NH2 + NH3). The second is a distal mechanism, where complete H-atom transfer at the distal N-atom to liberate an NH3 equivalent precedes transfers to the proximal N-atom (e.g., Fe-N2 + 3 e- + 3 H+ ? Fe?N + NH3). We also explore a new hybrid mechanism that first invokes a distal intermediate (Fe=NNH2) that then crosses to an alternating intermediate (Fe-N2H4) before releasing the first NH3 equivalent. We will use synthetic model complexes to test the viability of each of these mechanistic pathways, and to understand how the nuclearity, local geometry, and electronic structure of Fe- NxHy species control their relative stabilities and reactivity patterns. This knowledge will be applied to the study, via spectroscopic, electrochemical, and theoretical methods, of the first examples of single-site iron catalysts for N2-to-NH3 conversion that we discovered in the previous grant period, and towards the design of new N2-fixing catalysts with enhanced efficiency. Regardless of the precise mechanism for nitrogen reduction at the FeMoco, its ultimate solution will require comparison of spectroscopic data from the cofactor to related data obtained for well-defined model complexes. We will therefore continue to collaborate with researchers that specialize in spectroscopic studies of the FeMoco to make such comparisons, and other investigators with expertise complementary to our own. In sum, the functional Fe-NxHy model chemistry proposed will continue to play a critical role alongside current biochemical, spectroscopic, and theoretical model studies aimed at unraveling the chemical mechanism of biological nitrogen fixation.